Serotonin (5-HT) is a potent neuromodulator that regulates the spinal cord motoneurons, interneurons and sensory afferents involved in controlling posture and movement. 5-HT facilitates motoneuron excitability in part by activating Gq-protein-coupled 5-HT2 receptors that facilitate persistent calcium and sodium currents (persistent inward currents, or PICs), which in turn markedly amplify and prolong the motoneuron response to synaptic inputs. Because 5-HT arises mostly from the brainstem, spinal cord injury dramatically reduces 5-HT levels below the injury, and thus, immediately after injury PICs become small and motor output is accordingly severely weakened. However, the PICs undergo a remarkable recovery over the ensuing months after injury, becoming very large, despite the continued absence of most 5-HT below the injury. These large PICs play an important role in the recovery of residual motor function after chronic injury, and at the same time cause excess motoneuron activity that is manifested as severe muscle spasms. Thus, understanding what causes this spontaneous recovery of PICs and general motoneuron excitability is both important for understanding recovery of motor function and treating muscle spasms, the focus of our research. This proposal investigates the novel idea that the recovery of motoneuron PICs after chronic spinal cord injury may result from 5-HT2 receptors that become constitutively active, that is, spontaneously entering into their active state and initiating Gq-protein-mediated signaling without binding of 5-HT (Aim 1). In contrast, other 5-HT receptors that also modulate spinal cord excitability may not change with chronic spinal cord injury, and this is investigated in Aims 2 and 3 as follows. Normally, 5-HT regulates dorsal horn interneurons and associated sensory afferent transmission via Gi-coupled 5HT1 receptors. Loss of 5-HT with spinal cord injury causes an immediate loss of inhibition over afferent transmission, but this transmission is not restored to normal with chronic injury, suggesting that the 5-HT1 receptors do not compensate for lost 5-HT with constitutive activity. 5-HT also regulates the motoneuron membrane resting potential via Gs-coupled 5-HT6 (and 5-HT7) receptors, and likewise , membrane potential does not change with chronic, compared to acute, spinal cord injury, suggesting again that these 5-HT6 receptors do not become constitutively active with chronic injury. The final experiments (of Aim 4) examine the potential clinical application of Aims 1 - 3. These experiments explore whether drugs that modulate 5-HT2, 5-HT1 and 5-HT6 receptor activity (and associated PICs, afferent transmission and resting potential) can be used to help restore near-normal levels of postural tone and reflex patterns, while reducing muscle spasms. In summary, constitutive receptor activity provides an entirely new mechanism to explain spontaneous recovery of motor function after injury. Modulation of this constitutive receptor activity (or lack of activity) provides a novel approach to treating spasticity while restoring lost motor function. PUBLIC HEALTH RELEVANCE: The excitability of spinal neurons is highly dependent on the serotonin (5HT) released by axons originating in the brainstem and thus loss of 5HT in spinal injury has devastating consequences. Yet spinal neurons undergo remarkable adaptations post injury that in part tend to compensate for injury, despite the continued lack of 5HT itself. In this proposal we investigate whether this compensation occurs because 5HT receptors develop constitutive activity (activity without binding 5HT); this idea may provide an entirely new strategy for restoration of normal motor function in chronic spinal injury.